摘要
目的诱导表达并纯化细粒棘球绦虫EgAgB8/1重组抗原和EgAgB8/1-EgAgB8/2重组嵌合抗原,比较两个重组蛋白对囊型包虫病(CE)的血清学诊断价值。方法 IPTG诱导转染至E.coliBL21(DE3)LysS的pET32a-EgAgB8/1和pET32a-EgAgB8/1-EgAgB8/2重组原核表达质粒,表达和纯化EgAgB8/1重组蛋白和EgAgB8/1-EgAgB8/2重组嵌合蛋白,用SDS-PAGE电泳分析鉴定重组蛋白,并通过超声裂解法进行纯化,以CE病人及囊虫病人血清为一抗,Western blot法检测两个重组蛋白免疫反应性。结果细粒棘球绦虫重组抗原pET32a-EgAgB8/1和pET32a-EgAgB8/1-EgAgB8/2重组原核表达质粒得到成功诱导表达,经SDS-PAGE电泳分析,重组蛋白分子质量单位为28 ku和38 ku;Western blot结果表明,EgAgB8/1重组蛋白和EgAgB8/1-EgAgB8/2重组嵌合蛋白均能被CE病人血清特异性识别,16份CE病人血清中,以EgAgB8/1重组蛋白为抗原时11份阳性,以EgAgB8/1-EgAgB8/2重组蛋白为抗原时13份阳性;用EgAgB8/1和EgAgB8/1-EgAgB8/2重组蛋白检测12份囊虫病人血清,分别有5份和3份阳性。结论 EgAgB8/1-EgAgB8/2重组嵌合蛋白具有抗原特异性,对囊型包虫病的血清学诊断价值优于EgAgB8/1重组蛋白。
Objective To express and serologically evaluate the recombinant protein EgAgB8/1 and the recombinant chi meric protein EgAgB8/1-EgAgB8/2 from Echinococcus granulosus in order to serologically diagnose cystic echinococcus （CE）. Methods The recombinant prokaryotic expression vectors pET32a-EgAgB8/1 and pET32a-EgAgB8/1-EgAgB8/ 2 were introduced into E. coli BL21（DE3） LysS and induced with IPTG to express protein. The recombinant proteins obtained were purified by uhrasonication. The serological reactivity of these recombinant proteins was evaluated with West- ern blot using serum samples from patients with hydatidosis and cystieercosis as primary antibodies. Results E. granulosus recombinant antigens pET32a-EgAgB8/1 and pET32a EgAgB8/1-EgAgB8/2 were successfully expressed in a pro karyotic expression system. SDS-PAGE analysis revealed that the molecular mass of the recombinant proteins was 28 ku and 38 ku, respectively. Western blotting revealed that these recombinant proteins reacted specifically with serum samples from patients with hydatidosis. To determine the serological reaction of these recombinant proteins, the proteins were al lowed to react with 16 serum samples from patients with hydatidosis. Eleven samples reacted positively to the recombinant protein EgAgB8/1 and 13 reacted positively to the recombinant chimeric protein EgAgB8/1 EgAgB8/2. The proteins were also allowed to react with 12 serum samples from patients with cysticercosis. Five serum samples reacted positively to the recombinant protein EgAgB8/1 and 3 reacted positively to the recombinant chimeric protein EgAgB8/1-EgAgB8/2. Conclusion The recombinant chimeric protein EgAgB8/1-EgAgB8/2 possesses specific antigenicity and performs better than the recombinant protein EgAgBS/1 in the serological diagnosis of cystic echinococcosis.
出处
《中国病原生物学杂志》
CSCD
2011年第6期428-431,419,共5页
Journal of Pathogen Biology
基金
国家自然科学基金项目(No.30760229)
新疆维吾尔自治区高校科研计划项目(No.XJEDU2008S31)
